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Substance P and Inflammatory Pain: Getting It Wrong and Right Simultaneously

2019; Cell Press; Volume: 101; Issue: 3 Linguagem: Inglês

10.1016/j.neuron.2019.01.034

1097-4199

Edita Navratilova, Frank Porreca,

Coagulation, Bradykinin, Polyphosphates, and Angioedema

How do neuropeptides participate in the classic neuroinflammatory sequelae of tissue injury that includes pain, immune cell infiltration, and swelling? In this issue of Neuron, Green et al., 2019Green D.P. Limjunyawong N. Gour N. Pundir P. Dong X. A mast-cell-specific receptor mediates neurogenic inflammation and pain.Neuron. 2019; 101 (this issue): 412-420Abstract Full Text Full Text PDF PubMed Scopus (160) Google Scholar reveal that MrgprB2/MrgprX2 is a mast cell substance-P-specific receptor that critically links neural and immune systems and offers new approaches for neuroinflammatory therapeutics. How do neuropeptides participate in the classic neuroinflammatory sequelae of tissue injury that includes pain, immune cell infiltration, and swelling? In this issue of Neuron, Green et al., 2019Green D.P. Limjunyawong N. Gour N. Pundir P. Dong X. A mast-cell-specific receptor mediates neurogenic inflammation and pain.Neuron. 2019; 101 (this issue): 412-420Abstract Full Text Full Text PDF PubMed Scopus (160) Google Scholar reveal that MrgprB2/MrgprX2 is a mast cell substance-P-specific receptor that critically links neural and immune systems and offers new approaches for neuroinflammatory therapeutics. Mankind has relied on the pain-relieving actions of opioids for centuries. Nevertheless, their undesirable effects have long been recognized, and the need for new pain therapies remains a medical priority. Despite significant increases in the understanding of pain neurobiology, new classes of drugs for the treatment of pain have been slow to emerge (Borsook et al., 2014Borsook D. Hargreaves R. Bountra C. Porreca F. Lost but making progress--where will new analgesic drugs come from?.Sci. Transl. Med. 2014; 6: 249sr3Crossref PubMed Scopus (74) Google Scholar, Woolf, 2010Woolf C.J. Overcoming obstacles to developing new analgesics.Nat. Med. 2010; 16: 1241-1247Crossref PubMed Scopus (162) Google Scholar), resulting in a loss of confidence about the translational relevance of preclinical insights of nociceptive mechanisms to the human experience of pain. An example often cited to demonstrate the lack of translation between preclinical studies of pain and efficacy in humans is the experience from development of antagonists of the neurokinin 1 (NK1) receptor (Borsook et al., 2012Borsook D. Upadhyay J. Klimas M. Schwarz A.J. Coimbra A. Baumgartner R. George E. Potter W.Z. Large T. Bleakman D. et al.Decision-making using fMRI in clinical drug development: revisiting NK-1 receptor antagonists for pain.Drug Discov. Today. 2012; 17: 964-973Crossref PubMed Scopus (46) Google Scholar). The recent work of Prof. Xinzhong Dong and his colleagues reported in this issue of Neuron (Green et al., 2019Green D.P. Limjunyawong N. Gour N. Pundir P. Dong X. A mast-cell-specific receptor mediates neurogenic inflammation and pain.Neuron. 2019; 101 (this issue): 412-420Abstract Full Text Full Text PDF PubMed Scopus (160) Google Scholar) now begins to shed new light on this issue. The physiological role of nociceptors is to convey warning of actual or impending tissue damage to the organism. In humans, activation of nociceptors produces pain, while local anesthetic inactivation or genetically-based dysfunction (Bennett and Woods, 2014Bennett D.L. Woods C.G. Painful and painless channelopathies.Lancet Neurol. 2014; 13: 587-599Abstract Full Text Full Text PDF PubMed Scopus (218) Google Scholar) of these fibers can prevent the perception of pain. Nociceptors contribute to pain not only by afferent transmission of information to the spinal dorsal horn but also through their efferent functions. Damage to tissues produces pain that underlies immediate protective behaviors as well as inflammation in which infiltration of immune cells to the injury site clears cellular debris and promotes wound healing. The signature of inflammation is the “tetrad response” of dolor (pain), rubor (redness), calor (heat), and tumor (swelling). Inherent in the infiltration of immune cells to the injury site and to these localized responses is the axon reflex, a process in which neurotransmitters are released locally by terminals of damaged or irritated axons and their branches. Neuropeptides, along with other pronociceptive mediators often described as an “inflammatory soup,” play a key role in the tetrad response. While the peripheral consequences of neuropeptides released from primary afferents in the setting of tissue injury were appreciated, the development of NK1 antagonists for pain therapy was largely based on the concept of interrupting neurotransmission of substance P from primary afferent nociceptors to central pain pathways in the spinal dorsal horn and more rostrally (Mantyh, 2002Mantyh P.W. Neurobiology of substance P and the NK1 receptor.J. Clin. Psychiatry. 2002; 63: 6-10PubMed Google Scholar). Substantial evidence supports a role of substance P in promoting pain. In preclinical models, intrathecal or local peripheral injection of substance P elicits pain behaviors (Partridge et al., 1998Partridge B.J. Chaplan S.R. Sakamoto E. Yaksh T.L. Characterization of the effects of gabapentin and 3-isobutyl-gamma-aminobutyric acid on substance P-induced thermal hyperalgesia.Anesthesiology. 1998; 88: 196-205Crossref PubMed Scopus (148) Google Scholar). NK1 receptors are found postsynaptically in the spinal dorsal horn but also on immune cells, smooth muscle, blood vessels, and other cells in the periphery (Schäffer et al., 1998Schäffer M. Beiter T. Becker H.D. Hunt T.K. Neuropeptides: mediators of inflammation and tissue repair?.Arch. Surg. 1998; 133: 1107-1116Crossref PubMed Scopus (154) Google Scholar) where release of substance P promotes neurogenic inflammation (Pedersen-Bjergaard et al., 1991Pedersen-Bjergaard U. Nielsen L.B. Jensen K. Edvinsson L. Jansen I. Olesen J. Calcitonin gene-related peptide, neurokinin A and substance P: effects on nociception and neurogenic inflammation in human skin and temporal muscle.Peptides. 1991; 12: 333-337Crossref PubMed Scopus (118) Google Scholar). The pharmaceutical industry invested heavily in the discovery and development of NK1 receptor antagonists as therapies for pain, but clinical evaluation of these molecules demonstrated no significant analgesic effects (Borsook et al., 2012Borsook D. Upadhyay J. Klimas M. Schwarz A.J. Coimbra A. Baumgartner R. George E. Potter W.Z. Large T. Bleakman D. et al.Decision-making using fMRI in clinical drug development: revisiting NK-1 receptor antagonists for pain.Drug Discov. Today. 2012; 17: 964-973Crossref PubMed Scopus (46) Google Scholar). The failure of NK1 antagonists to produce analgesia reverberated across the pain community, raising doubts about the predictive validity of preclinical studies. Addressing the question of translation directly was complicated by the structural differences between the human and rodent NK1 receptors so that the NK1 antagonists optimized for human use were of limited utility for preclinical studies in commonly used and well-characterized models of pain. In retrospect, it seems reasonable that blocking the actions of substance P at the spinal level might not be sufficiently effective given the numerous other transmitters in primary afferents that promote nociceptive transmission. On the other hand, given the known role of substance P in neurogenic inflammation, it was puzzling that NK1 antagonists would not have significant anti-inflammatory actions at peripheral sites. Clinically available NK1 antagonists are effective for emesis but are not used as anti-inflammatory agents. In a landmark study published in 2015 (McNeil et al., 2015McNeil B.D. Pundir P. Meeker S. Han L. Undem B.J. Kulka M. Dong X. Identification of a mast-cell-specific receptor crucial for pseudo-allergic drug reactions.Nature. 2015; 519: 237-241Crossref PubMed Scopus (735) Google Scholar), Dong and colleagues demonstrated that the Mas-related G-protein-coupled receptor MrgprB2, as well as its human homolog MrgprX2, was activated by substance P. These findings raised the possibility that blockade of the NK1 receptor was insufficient to prevent some of the local pronociceptive and proinflammatory actions of substance P that were associated with tissue injury. In the current study, Green et al., 2019Green D.P. Limjunyawong N. Gour N. Pundir P. Dong X. A mast-cell-specific receptor mediates neurogenic inflammation and pain.Neuron. 2019; 101 (this issue): 412-420Abstract Full Text Full Text PDF PubMed Scopus (160) Google Scholar have tested this possibility by determining whether substance P released from primary afferents can engage the MrgprB2 receptor on mast cells to promote the release of inflammatory mediators and to increase the infiltration to the injury site of immune cells, including lymphocytes, neutrophils, and monocytes. The mast cell is a key link between the innate immune response and the nervous system. Green et al., 2019Green D.P. Limjunyawong N. Gour N. Pundir P. Dong X. A mast-cell-specific receptor mediates neurogenic inflammation and pain.Neuron. 2019; 101 (this issue): 412-420Abstract Full Text Full Text PDF PubMed Scopus (160) Google Scholar performed a series of elegant studies to demonstrate that in the absence of mast cells or activation of the MrgprB2 receptor, the consequences of inflammatory pain induced by incisional injury (or complete Freund’s adjuvant [CFA]) were greatly reduced. Critically, they confirmed important aspects of these findings with the human homolog of the mouse receptor, the MrgprX2. Green et al., 2019Green D.P. Limjunyawong N. Gour N. Pundir P. Dong X. A mast-cell-specific receptor mediates neurogenic inflammation and pain.Neuron. 2019; 101 (this issue): 412-420Abstract Full Text Full Text PDF PubMed Scopus (160) Google Scholar confirmed that the MrgprB2 receptor was expressed exclusively on mast cells and not on dorsal root ganglion (DRG) neurons and proceeded to demonstrate that mice lacking the MrgprB2 receptor showed lower thermal and mechanical hypersensitivity (dolor) following incision (or CFA) injury that was accompanied by a decrease in expression of injury-related ATF3 in DRG cells when compared to wild-type (WT) controls. These findings linked the MrgprB2 receptor on mast cells with pain and neuronal activation. Ablation of mast cells with diphtheria toxin in transgenic mice expressing the human diphtheria toxin receptor in mast cells (MrgprB2Cre × Cre-dependent ROSA26DTR) similarly showed reductions in inflammatory pain. Following injury, MrgprB2−/− mice also had significantly decreased swelling (tumor) than controls. Using flow cytometry, Green et al., 2019Green D.P. Limjunyawong N. Gour N. Pundir P. Dong X. A mast-cell-specific receptor mediates neurogenic inflammation and pain.Neuron. 2019; 101 (this issue): 412-420Abstract Full Text Full Text PDF PubMed Scopus (160) Google Scholar then demonstrated that MrgprB2−/− mice had lower increases in hindpaw mast cells compared to WT mice despite equivalent numbers at baseline. These findings led to the hypothesis that activation of the MrgprB2 receptors may mediate the recruitment of immune cells to the injury site. Green et al., 2019Green D.P. Limjunyawong N. Gour N. Pundir P. Dong X. A mast-cell-specific receptor mediates neurogenic inflammation and pain.Neuron. 2019; 101 (this issue): 412-420Abstract Full Text Full Text PDF PubMed Scopus (160) Google Scholar again used flow cytometry in hindpaw biopsies of MrgprB2−/− and WT mice to demonstrate decreased infiltration of leukocytes (CD45), neutrophils ((Ly)6G+CD11b+), and monocytes (CD11b+Ly6G−). These findings provided strong support for a critical role of the MrgprB2 receptor in pain, swelling, and infiltration of immune cells, but the transmitter mediating these responses remained to be identified. Based on their previous work (McNeil et al., 2015McNeil B.D. Pundir P. Meeker S. Han L. Undem B.J. Kulka M. Dong X. Identification of a mast-cell-specific receptor crucial for pseudo-allergic drug reactions.Nature. 2015; 519: 237-241Crossref PubMed Scopus (735) Google Scholar), Green et al., 2019Green D.P. Limjunyawong N. Gour N. Pundir P. Dong X. A mast-cell-specific receptor mediates neurogenic inflammation and pain.Neuron. 2019; 101 (this issue): 412-420Abstract Full Text Full Text PDF PubMed Scopus (160) Google Scholar evaluated whether substance P, an agonist at the MrgprB2 receptor, could recruit immune cells. As the NK1 receptor is the canonical receptor for substance P, they generated NK1−/− mice, confirmed the expression of this receptor to the spinal dorsal horn but not in the DRG or mast cells in WT mice, and then demonstrated no significant differences in the recruitment of immune cells to the injury site between NK1−/− and WT mice. To test the role of substance P directly in the recruitment of immune cells and pain, Green et al., 2019Green D.P. Limjunyawong N. Gour N. Pundir P. Dong X. A mast-cell-specific receptor mediates neurogenic inflammation and pain.Neuron. 2019; 101 (this issue): 412-420Abstract Full Text Full Text PDF PubMed Scopus (160) Google Scholar then used an immunoneutralization strategy with antibody against the peptide. They demonstrated that sequestration of substance P with a specific antibody, but not an isotype control peptide, prior to surgery was able to decrease infiltrating immune cells and paw swelling, directly implicating this peptide, but not the NK1 receptor, in incisional-induced inflammation. The recruitment of immune cells to the injury site therefore seems to require substance-P-induced activation of the MrgprB2 receptor on mast cells, presumably through the release of inflammatory cytokines and chemokines (Figure 1). Green et al., 2019Green D.P. Limjunyawong N. Gour N. Pundir P. Dong X. A mast-cell-specific receptor mediates neurogenic inflammation and pain.Neuron. 2019; 101 (this issue): 412-420Abstract Full Text Full Text PDF PubMed Scopus (160) Google Scholar pursued this question using a human mast cell line, the LAD2 cells that express the human homolog receptor MrgprX2. Following application of substance P to this cell line, they evaluated the release of 12 cytokines and 11 chemokines and found significant increases only in TNFα, GM-CSF, and IL8, along with CCL2, CCL3, and CCL4. The substance-P-induced increases in the levels of these cytokines and chemokines was prevented by siRNA directed at the MrgprX2 receptor in the LAD2 cells establishing the direct role of substance P and the MrgprX2 receptor in releasing these mediators in a human cell line. They then confirmed these results by demonstrating that incisional injury similarly increased the release of these mediators in mice and that this effect was diminished in MrgprB2−/− mice. A major finding of these studies is the identification of MrgprB2 and its human homolog MrgprX2 as novel molecular targets on mast cells that underlie neuropeptide-induced inflammatory responses including pain, swelling, and infiltration of immune cells to the injury site. The contribution of mast cells to inflammatory processes and pain is well known. Neuropeptides, including substance P, can provoke mast cells to release pro-inflammatory mediators, including TNFα, GM-CSF, IL8, CCL2, CCL3, and CCL4, cytokines, and chemokines, that alter vascular permeability and to promote pain in both animals and humans. CCL3 may directly sensitize the TRPV1 channel on peripheral nociceptors, and CCL4 has been demonstrated to be upregulated following peripheral nerve injury. The current findings now demonstrate that this effect is mediated by actions of substance P on the MrgprB2/X2 receptor and not on the NK1 receptor. A key concept in the development of NK1 antagonists as novel pain therapeutics was that pain could be treated by targeting transmission of nociceptive information at the first synapse in the spinal dorsal horn, avoiding the unwanted side effects associated with engagement of opioid receptors in the brain. While this approach remains attractive and the basis of many present-day drug discovery efforts (e.g., Nav1.7 blockers), potential safety concerns related to the blockade of the physiological role of pain in preventing harm persist. The axon reflex is activated physiologically in the setting of injury and promotes the activation and sensitization of nociceptors through both neural and immune mechanisms that are linked through engagement of mast cells. Data from the study of Green et al., 2019Green D.P. Limjunyawong N. Gour N. Pundir P. Dong X. A mast-cell-specific receptor mediates neurogenic inflammation and pain.Neuron. 2019; 101 (this issue): 412-420Abstract Full Text Full Text PDF PubMed Scopus (160) Google Scholar demonstrate that preventing the activation of mast cells from released substance P by blocking the MrgprB2 receptor or by ablation of these cells greatly diminishes, but does not completely abolish, pain responses. Prevention of nociceptor sensitization is a validated mechanism by which analgesia can be elicited in humans with non-steroidal anti-inflammatory drugs (NSAIDs). Additionally, neurosteroids are drugs that produce powerful suppression of the immune response and can be very effective in producing pain relief in many patients. Neuroinflammatory processes likely generate the most common types of pain in humans, including conditions such as musculoskeletal and visceral pains, and many chronic pain syndromes, such as neuropathic and cancer pains, likely include inflammatory components. Therefore, blocking these processes through a novel mechanism that involves inflammatory mediators that may not be targeted by currently available therapies, and without a generalized suppression of the immune response, may increase the numbers of patients that can be effectively treated. The activation of MrgprX2, the human receptor, in a mast cell line by substance P and the release of cytokines and chemokines known to promote neuroinflammatory processes demonstrate translation between preclinical and human models. These studies also help us to understand the puzzling result that blocking substance P actions with NK1 antagonists has no apparent effects on neuroinflammation in humans despite a clear proinflammatory role of this peptide. The studies of Dong and colleagues suggest that while we may have been right in detecting the prominent role of substance P in pain and inflammation, perhaps we targeted the wrong receptor. Future work will determine whether targeting the MrgprX2 receptor will lead to effective treatments of neuroinflammatory pain without the unwanted effects of many of our currently available therapeutics. A Mast-Cell-Specific Receptor Mediates Neurogenic Inflammation and PainGreen et al.NeuronJanuary 24, 2019In BriefGreen et al. show that activation of the mast cell receptor Mrgprb2/X2 by the neuropeptide substance P leads to cytokine release and recruitment of immune cells contributing to inflammatory pain. Full-Text PDF Open Archive